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1.
BioPharm International ; 35(2):26-29, 2022.
Article in English | Scopus | ID: covidwho-2012042
2.
Frontiers in Immunology ; 13, 2022.
Article in English | EMBASE | ID: covidwho-2009868

ABSTRACT

Extracellular vesicles (EVs) are membrane-bound particles released by cells in various (patho)physiological conditions. EVs can transfer effector molecules and elicit potent responses in recipient cells, making them attractive therapeutic agents and drug delivery platforms. In contrast to their tremendous potential, only a few EV-based therapies and drug delivery have been approved for clinical use, which is largely attributed to limited therapeutic loading technologies and efficiency. As EV cargo has major influence on their functionality, understanding and translating the biology underlying the packaging and transferring of biomolecule cargos (e.g. miRNAs, pathogen antigens, small molecule drugs) into EVs is key in harnessing their therapeutic potential. In this review, through recent insights into EVs’ content packaging, we discuss different mechanisms utilized by EVs during cargo packaging, and how one might therapeutically exploit this process. Apart from the well-characterized EVs like exosomes and microvesicles, we also cover the less-studied and other EV subtypes like apoptotic bodies, large oncosomes, bacterial outer membrane vesicles, and migrasomes to highlight therapeutically-diverse opportunities of EV armoury.

3.
Current Nanoscience ; 18(6):717-725, 2022.
Article in English | EMBASE | ID: covidwho-2005811

ABSTRACT

Background: The spread of new coronavirus 2019, the causative agent of viral pneumonia documented in Wuhan, brought a recent public health crisis globally. The best solution to overcome this pandemic is developing suitable and effective vaccines and therapeutics. However, discovering and creating a new drug is a lengthy process requiring rigorous testing and validation. Objective: Despite many newly discovered and old repurposed COVID-19 drugs under clinical trial, more emphasis should be given to research on COVID-19 NPs-based medicines, which could improve the efficacy of antiviral drugs to reduce their side effects. The use of NPs as carriers can reduce the frequency and duration of drug ingestion, enhance approved antiviral therapeutics' effectiveness, and overcome their limitations, such as low bioavailability. Besides, they can play a crucial role in fighting against the COVID-19 pandemic. In this regard, nanotechnology provides opportunities to develop new strategies for preventing, diagnosing, and treating COVID-19. Conclusion: This review highlighted the importance of NMs-based technical solutions in antiviral drugs for testing against the SARS-CoV-2 virus emergencies in the form of nanotherapeutics.

4.
Journal of Drug Delivery Science and Technology ; 74, 2022.
Article in English | EMBASE | ID: covidwho-1996814
5.
Frontiers in Microbiology ; 13, 2022.
Article in English | EMBASE | ID: covidwho-1987517

ABSTRACT

Viral infections are a major cause of severe, fatal diseases worldwide. Recently, these infections have increased due to demanding contextual circumstances, such as environmental changes, increased migration of people and product distribution, rapid demographic changes, and outbreaks of novel viruses, including the COVID-19 outbreak. Internal variables that influence viral immunity have received attention along with these external causes to avert such novel viral outbreaks. The gastrointestinal microbiome (GIM), particularly the present probiotics, plays a vital role in the host immune system by mediating host protective immunity and acting as an immune regulator. Bacteriocins possess numerous health benefits and exhibit antagonistic activity against enteric pathogens and immunobiotics, thereby inhibiting viral infections. Moreover, disrupting the homeostasis of the GIM/host immune system negatively affects viral immunity. The interactions between bacteriocins and infectious viruses, particularly in COVID-19, through improved host immunity and physiology are complex and have not yet been studied, although several studies have proven that bacteriocins influence the outcomes of viral infections. However, the complex transmission to the affected sites and siRNA defense against nuclease digestion lead to challenging clinical trials. Additionally, bacteriocins are well known for their biofunctional properties and underlying mechanisms in the treatment of bacterial and fungal infections. However, few studies have shown the role of probiotics-derived bacteriocin against viral infections. Thus, based on the results of the previous studies, this review lays out a road map for future studies on bacteriocins for treating viral infections.

6.
Cancer Research ; 82(12), 2022.
Article in English | EMBASE | ID: covidwho-1986485

ABSTRACT

Introduction: Despite the development of two mRNA vaccines, there is an urgent unmet need of finding new antiviral strategies. One such potential antiviral strategy is to target the synthetic lethal (SL) partners of transcriptionally altered genes in infected host cells, thereby selectively killing them to halt the infection at its heels (Mast FD, JCB, 2020). Methods: Here we conduct a first proof-of-concept SL inference approach to predict anti-SARS-CoV-2 targets in a systematic genome-wide manner. This effort capitalizes on our recently published pipeline for inferring clinically relevant SL interactions in cancer (Lee et al, Cell, 2021). Based on the latter, we comprehensively analyzed multiple in vitro and in vivo bulk and single-cell RNA-sequencing datasets of SARS-CoV-2 infection to predict candidate antiviral targets that are SL with altered host genes. Importantly, as our predictions are fine-tuned based on the analysis of patients' data, they are more likely to be of translational value. Results: Our key results are twofold:1) The predicted SL-based targets are highly enriched for genes that are reported in four SARS-CoV-2 CRISPR-Cas9 genome-wide genetic screens to inhibit growth of infected cells. 2) A subset of top predicted 26 genes were experimentally tested in a targeted siRNA screen conducted in both infected and non-infected human Caco-2 cells. Remarkably, as expected given that these targets were predicted to be SL specific with genes upregulated in infected cells, indeed, knocking down these targets reduced viral replication and cell viability only under the infected condition without harming non-infected cells. Conclusion: In summary, this study is the first to demonstrate the potential of a synthetic lethality approach to identify viral (specifically anti-SARS-CoV-2) targets. Importantly, as both single cell and bulk transcriptomics patients' data is considered from both infected people and controls, they are more likely to be of clinical relevance. Targeting host genes identified via an SL-based approach is probably more suitable when the infection is at the early stage and host can still tolerate the loss of infected host cells.

8.
American Journal of Respiratory and Critical Care Medicine ; 205(1), 2022.
Article in English | EMBASE | ID: covidwho-1927783

ABSTRACT

Rationale: Cell-penetrating peptides are able to cross membranes and deliver cargoes in a functional form. Our prior work identified a 12-amino acid, cardiac targeting peptide (APWHLSSQYSRT). Studies into its mechanism of transduction led to the identification of two lung targeting peptides (LTPs), S7A and R11A. Here we report on a) the comparative lung uptake of S7A versus R11A, b) complete biodistribution of R11A, c) show that cyclic versions are -100-fold more efficient than linear counterparts, d) uptake is via a non-endocytic pathway, and e) cyclic R11A's (cR11A) ability to deliver siRNA targeting structural proteins of SARS-CoV-2 and act as an anti-viral. Methods: Linear LTPs were synthesized with N-terminal labeled with Cyanine 5.5 (Cy5.5). Cyclic versions were synthesized with lysine added to the N-terminus, cyclized through a peptide bond, with a side NH-group labeled with Cy5.5. cR11A was conjugated to siRNA duplexes via a DTME linker. Wild-type, CD1 mice, were injected with S7A or R11A at 10, 5, and 1mg/Kg, peptides allowed to circulate for 15mins, mice euthanized, lung along with multiple other organs dissected and imaged using In Vivo Imaging Systems (IVIS, Perkin-Elmer) followed by confocal microscopy. CD1 mice were injected with R11A, 5mg/Kg, and euthanized at different time intervals for biodistribution studies. Endocytosis studies were done using serum-starved human bronchial epithelial cells (HBEC) incubated with fluorescently labeled transferrin and LTP-S7A or LTP- R11A. Lastly, anti-viral activity was tested in HBECs pre-treated with cR11A-siRNA followed by viral infection. Results: Mice injected with LTP-S7A or LTP-R11A showed robust uptake of the peptides by lung tissue, with R11A showing an increasingly favorable lung:liver ratio with decreasing dose. Lung uptake of R11A peaked at 120mins with complete dissipation of fluorescence by 24 hours. In Vitro studies in HBECs showed no co-localization of transferrin with LTPs, ruling out endocytosis as a mechanism of uptake. Comparison of linear versus cyclic peptides using FACS showed cyclic peptides had -100-fold increased transduction efficiency over their linear counterparts. cR11A conjugated to ant-spike, and anti-envelop proteins showed an anti-viral effect with EC90 of 0.6uM and 1.0μM, respectively. Conclusions: We have identified two novel lung-targeting peptides capable of acting as delivery vectors. Peak uptake of R11A occurred at 120mins. Furthermore, this uptake was not via endocytosis, and cyclic versions were -100-fold more efficiently taken up. Lastly, as proof of concept, we show cR11A acts as a vector and delivers siRNA to HBECs in a functional form, and act as anti-virals.

9.
American Journal of Respiratory and Critical Care Medicine ; 205(1), 2022.
Article in English | EMBASE | ID: covidwho-1927749

ABSTRACT

Introduction: Management of acute respiratory distress including COVID-19 pneumonia involves O2 supplementation, which is lifesaving, but causes severe hyperoxic acute lung injury (HALI). AT2 cells are the most affected cell type in hyperoxia (HO). NADPH oxidase (NOX) is a major source of reactive oxygen species (ROS) in HO. NOX4, the only functionally active NOX present in mitochondria, and primarily produces H2O2 as well as mtROS has been shown to be involved in several human pathologies. Not much is known about NOX4-induced mitochondrial injury in HALI. The current study aims to determine the role of AT2 epithelial cell NOX4 in HALI and the impact of HO on the modulation of mtROS and mitochondrial dynamics in HALI. Methods: Nox4-/-Spc-Cre animals were generated using tamoxifen induction and the knockdown was validated. The Nox4- /-Spc-Cre knockout (KO) and wild type (WT) mice were exposed to room air (NO) or 95% O2 (HO) for 66h to study the structural and functional changes in the lung. Transmission Electron Microscopy (TEM) was used to study the HO-induced changes in mitochondria. Isolated primary AT2 and/ mouse lung epithelial (MLE) cell line was investigated for mtROS, mt dynamics and apoptosis. Mitochondrial injury was assessed in Nox4 WT and Nox4 silenced cells. Results: C57BL/6J WT animals subjected to HO for 66h showed increased expression of NOX4, determining the role of NOX4 in HALI. The H&E staining demonstrated significant HALI in Nox4 WT animals exposed to HO compared to Nox4 KO as determined by increased infiltration of neutrophils, alveolar wall thickening and presence of proteinaceous debris in the alveolar space. Further, increased BAL cell count and protein levels, increased AT2 cell death and elevation of the proinflammatory cytokine IL- 6 and the chemokine KC was seen in WT animals compared to Nox4 KO. Analysis of lung tissues by TEM showed mitochondrial swelling, cristae damage and mitophagy in AT2 cells due to HO. Changes in mt injury markers were also observed. HO-induced NOX4 increase in primary AT2/ MLE-12 cells resulted in increased mtROS production and apoptosis, which was reduced with Nox4 siRNA silencing. Conclusion: This study suggests that the HO induced NOX4 expression in mouse lung, and deletion of Nox4 gene in AT2 cells reduced mtROS production and apoptosis and protected the lungs from severe hyperoxic lung injury. These results suggest NOX4 as a potential target for the treatment of HALI.

10.
Virologie ; 26(2):111, 2022.
Article in French | EMBASE | ID: covidwho-1913051

ABSTRACT

The interferon (IFN) response is a critical arm of the innate immune response and a major host defense mechanism against viral infections. Numerous genes that contribute to this antiviral state remain to be identified and characterized. Using large-scale loss-of-function strategies, we screened siRNAs or gRNAs libraries targeting hundreds of IFNstimulated genes (ISGs) in IFN-treated cells infected with human RNA viruses, including SARS-CoV-2, Zika virus or tick-borne encephalitis virus. We recovered previously unrecognized human genes able to modulate the replication of these RNA viruses in an IFN-dependent manner. For instance, we identified the chromatin remodeling protein MTA2 as a potent flavivirus-specific antiviral factor. Mechanistic studies to decipher the molecular mechanisms by which these novel antiviral genes are functioning are on-going. We are also expanding our studies to the identification and characterization of ISGs in animal species that serve as viral reservoirs, such as bats. Our work should open new perspectives to target weakness points in the life cycle of these emerging RNA viruses.

11.
Virologie ; 26(2):120, 2022.
Article in English | EMBASE | ID: covidwho-1912931

ABSTRACT

Establishment of the interferon (IFN)-mediated antiviral state provides a crucial initial line of defense against viral infection. Numerous genes that contribute to this antiviral state remain to be identified. Using a loss-of-function strategy, we screened an original library of 1156 siRNAs targeting 386 individual curated human genes in stimulated microglial cells infected with Zika virus (ZIKV), an emerging RNA virus that belongs to the flavivirus genus. The screen recovered twenty-one potential host proteins that modulate ZIKV replication in an IFN-dependent manner, including the previously known IFITM3 and LY6E. Further characterization contributed to delineate the spectrum of action of these genes towards other pathogenic RNA viruses, including Hepatitis C virus and SARS-CoV-2. Our data revealed that APOL3 acts as a proviral factor for ZIKV and several other related and unrelated RNA viruses. In addition, we showed that MTA2, a chromatin remodeling factor, possesses potent flavivirus-specific antiviral functions. We are currently investigating the molecular mechanisms behind IFN-dependent flaviviral restriction of MTA2. Our work identified previously unrecognized genes that modulate the replication of RNA viruses in an IFN-dependent way, opening new perspectives to target weakness points in the life cycle of these viruses.

12.
Topics in Antiviral Medicine ; 30(1 SUPPL):8, 2022.
Article in English | EMBASE | ID: covidwho-1880637

ABSTRACT

Background: We recently showed that genuine SARS-CoV-2 hijacks endogenously expressed interferon-induced transmembrane proteins, especially IFITM2, as entry cofactors for efficient infection (Prelli Bozzo, Nchioua et al., Nat. Com., 2021). This came as a surprise, since IFITMs have been reported to inhibit entry of numerous enveloped viruses, including SARS-CoV-2. However, most data were obtained using IFITM overexpression and pseudoparticle infection assays. In our initial study, we used a SARS-CoV-2 strain isolated in the Netherlands in February 2020 (NL-02-2020). Since then several "variants of concern" (VOCs) have emerged that show increased transmission fitness and evasion of vaccine-induced immunity. These VOCs contain various alterations in their Spike (S) proteins that may alter their dependency on entry cofactors. Here, we examined whether SARS-CoV-2 VOCs, including the currently dominating Delta variant, still depend on IFITMs for efficient infection and replication. Methods: To determine the role of IFITMs in infection of SARS-CoV-2 VOCs, we silenced IFITM1, 2, or 3 expression in Calu-3 cells using siRNAs and infected them with NL-02-2020 as well VOCs B.1.1.7, B.1.351, P.1 and B.1.617.2, also referred to as Alpha, Beta, Gamma and Delta variants, respectively. Viral entry and replication were quantified by qRT-PCR as well as TCID50 analysis. In addition, we determined the inhibitory effect of an α-IFITM2 antibody on VOC infection in iPSC-derived human alveolar epithelial type 2 (iAT2) cells. Results: Depletion of IFITM2 reduced viral RNA production from 31-(B.1.1.7) to 754-fold (P.1). In comparison, KD of IFITM1 generally had little effect, while silencing of IFITM3 resulted in 2-to 20-fold reduction of viral RNA yields by the four VOCs. An antibody directed against the N-terminus of IFITM2 inhibited SARS-CoV-2 VOC replication in iAT2 cells. Conclusion: Endogenously expressed IFITM proteins (especially IFITM2) are important cofactors for entry and replication of SARS-CoV-2 VOCs, including the Delta variant that currently dominates the COVID-19 pandemic.

13.
Topics in Antiviral Medicine ; 30(1 SUPPL):64, 2022.
Article in English | EMBASE | ID: covidwho-1880463

ABSTRACT

Background: SARS-CoV-2 primarily infects the lung but may also damage other organs including the brain, heart, kidney, and intestine. Central nervous system (CNS) disorders include loss of smell and taste, headache, delirium, acute psychosis, seizures, and stroke. Pathological loss of gray matter occurs in SARS-CoV-2 infection but it is unclear whether this is due to direct viral infection, indirect effects associated with systemic inflammation, or both. Methods: We used iPSC-derived brain organoids and primary human astrocytes from cerebral cortex to study direct SARS-CoV-2 infection, as confirmed by Spike and Nucleocapsid immunostaining and RT-qPCR. siRNAs, blocking antibodies, and small molecule inhibitors were used to assess SARS-CoV-2 receptor candidates. Bulk RNA-seq, DNA methylation seq, and Nanostring GeoMx digital spatial profiling were utilized to identify virus-induced changes in host gene expression. Results: Astrocytes were robustly infected by SARS-CoV-2 in brain organoids while neurons and neuroprogenitor cells supported only low-level infection. Based on siRNA knockdowns, Neuropilin-1, not ACE2, functioned as the primary receptor for SARS-CoV-2 in astrocytes. The endolysosomal two-pore channel protein, TPC, also facilitated infection likely through its regulatory effects on endocytosis. Other alternative receptors, including the AXL tyrosine kinase, CD147, and dipeptidyl protease 4 (DPP4), did not function as SARS-CoV-2 receptors in astrocytes. SARS-CoV-2 infection dynamically induced type I, II, and III interferons, and genes involved in Toll-like receptor signaling, MDA5 and RIG-I sensing of double-stranded RNA, and production of inflammatory cytokines. Genes activating apoptosis were also increased. Down-regulated genes included those involved in water, ion and lipid transport, synaptic transmission, and formation of cell junctions. Epigenetic analyses revealed transcriptional changes related to DNA methylation states, particularly decreased DNA methylation in interferon-related genes. Long-term viral infection of brain organoids resulted in progressive neuronal degeneration and death. Conclusion: Our findings support a model where SARS-CoV-2 infection of astrocytes produces a panoply of changes in the expression of genes regulating innate immune signaling and inflammatory responses. Deregulation of these genes in astrocytes produces a microenvironment within the CNS that ultimately disrupts normal neuron function, promoting neuronal cell death and CNS deficits.

14.
Topics in Antiviral Medicine ; 30(1 SUPPL):67, 2022.
Article in English | EMBASE | ID: covidwho-1880292

ABSTRACT

Background: Human immunodeficiency virus (HIV) and Influenza A virus (IAV) remain a global health concern. Further, emergence of novel coronavirus SARS-CoV-2, which rapidly became global pandemic, increases the concern in biomedical research field for antiviral treatment. To develop new antiviral therapy, we must need to understand the molecular and cellular mechanisms involved in assembly and replication. It is known for some viruses (HIV and IAV) that the host actin cytoskeleton has been involved in various stages of the virus life cycle. Regulation of actin cytoskeleton requires several actin binding proteins, which organize the actin filaments (F-actin) into higher order structures such as actin bundles, branches, filopodia and microvilli, for further assistance in viral particle production. Thus, our objective for this work is to understand the role of these actin regulator proteins, like cofilin and one of its cofactor WDR1, in viral particle assembly and release. Methods: Here we used a combination of different experimental methods like RNA interference, immunoblot, immunoprecipitation, immunofluorescence coupled to confocal and STED fluorescence microscopy. In order to study only virus release, and bypass viral entry, we set up a minimal system for virus-like particles production in transfected cells, giving HIV-1 Gag-VLP, Influenza M1-VLP and SARS-CoV-2 MNE-VLP (developed by D. Muriaux lab). For image analysis, we used Image J software. Statistical analysis was performed with non-parametric t-tests or one-way Anova test. Results: Using siRNA strategy, we have shown that upon knock down of actin protein cofilin or WDR1, HIV-1 and IAV particles production increases in contrario to SARS-CoV-2 VLP release. Further, using immunoprecipitation, we report that HIV-1 Gag is able to form an intracellular complex with WDR1 and cofilin. Similarly, IAV-M1, which like HIV Gag-MA binds with plasma membrane phospholipids, is able to form an intracellular complex with cofilin. These results suggested that virus budding from the host cell plasma membrane seemed restricted by the cofilin/WDR1 complex. Finally, using confocal/STED microscopy on cell producing VLP, we observed actin fibers rearrangement with cell protrusions, suggesting a role for actin in viral particles assembly and release. Conclusion: In conclusion, regulators of actin dynamic are involved in HIV-1 Gag, IAV-M1 and SARS-CoV-2 VLP production but play a differential role in assembly and release of these RNA enveloped viruses.

15.
Topics in Antiviral Medicine ; 30(1 SUPPL):67, 2022.
Article in English | EMBASE | ID: covidwho-1879932

ABSTRACT

Background: A promising approach to tackle the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) could be small interfering (si)RNAs. However, it is unclear so far, which viral replication steps can be efficiently inhibited with siRNAs. Here we report the first-ever in-depth analysis of RNAi-accessible SARS-CoV-2 replication steps. Methods: siRNAs were designed against four genomic regions of SARS-CoV-2. Initial screening of siRNA activity was performed with a dual luciferase reporter assay. Efficacy of siRNAs to terminate various viral replication steps was analyzed by infecting VeroE6 cells with wildtype SARS-CoV-2 or a GFP expressing recombinant SARS-CoV-2 and monitoring viral spread in real-time by time-lapse fluorescence microscopy. Positive and negative sense viral RNA transcripts were distinctly quantified via sense specific cDNA synthesis and reverse-transcriptase quantitative PCR. Finally, the antiviral activity of the siRNAs was primarily evaluated in a highly relevant model, SARS-CoV-2 infected human lung explants. Results: When applied in a prophylactic fashion, siRNAs were able to target genomic RNA (gRNA) of SARS-CoV-2 after cell entry, terminating replication before start of transcription, thereby preventing cytopathic effects. Surprisingly, siRNAs were not active against intermediate negative sense transcripts formed during replication. Targeting sequences that are commonly shared by all viral transcripts indeed allowed a simultaneous suppression of gRNA and subgenomic (sg)RNAs by a single siRNA. However, siRNAs that targeted ORF1 which is solely part of gRNA, presented an enhanced antiviral activity. We show that the reason for this was that siRNAs that targeted the common regions of transcripts were outcompeted by the highly abundant sgRNAs. Based on these findings, we developed a chemically stabilized siRNA, which targets a highly conserved region of ORF1, and which inhibited SARS-CoV-2 replication by >90% ex vivo in explants of the human lung. Conclusion: Our work strongly encourages the development of siRNA-based therapies for COVID-19 and suggests that early therapy start, or prophylactic application, together with targeting ORF1, might be key for high antiviral efficacy.

16.
Front Oncol ; 12: 891812, 2022.
Article in English | MEDLINE | ID: covidwho-1862637

ABSTRACT

Despite the early promise of RNA therapeutics as a magic bullet to modulate aberrant signaling in cancer, this field remains a work-in-progress. Nevertheless, RNA therapeutics is now a reality for the treatment of viral diseases (COVID-19) and offers great promise for cancer. This review paper specifically investigates RNAi as a therapeutic option for HCC and discusses a range of RNAi technology including anti-sense oligonucleotides (ASOs), Aptamers, small interfering RNA (siRNA), ribozymes, riboswitches and CRISPR/Cas9 technology. The use of these RNAi based interventions is specifically outlined in three primary strategies, namely, repressing angiogenesis, the suppression of cell proliferation and the promotion of apoptosis. We also discuss some of the inherent chemical and delivery problems, as well as targeting issues and immunogenic reaction to RNAi interventions.

17.
Tissue Engineering - Part A ; 28(SUPPL 1):S649, 2022.
Article in English | EMBASE | ID: covidwho-1852886

ABSTRACT

The COVID-19 pandemic has shown how revolutionary treatments based on gene therapeutics has helped overcome a once-in-acentury pandemic and has given new momentum to gene therapy research for a myriad of applications. The field of regenerative medicine is well placed to be a beneficiary whereby, for example, gene therapy might be a valuable tool to avoid the limitations of local delivery of growth factors. While non-viral vectors are typically inefficient at transfecting cells, our group have had significant success in this area using a scaffold-mediated gene therapy approach for regenerative applications[1, 2]. These gene activated scaffold platforms not only act as a template for cell infiltration and tissue formation, but also can be engineered to direct autologous host cells to take up specific genes and then produce therapeutic proteins in a sustained but eventually transient fashion. Similarly, we have demonstrated how scaffold-mediated delivery of siRNAs[3] and miRNA[4, 5] can be used to silence specific genes associated with reduced repair or pathological states. This presentation will provide an overview of ongoing research in our lab in this area with a particular focus on gene-activated biomaterials for promoting bone, cartilage, nerve and wound repair. Focus will also be placed on advances we are making in using 3D printing of gene activated bioinks to produce next generation medical devices for tissue repair.

18.
Journal of Excipients and Food Chemicals ; 13(1):4-17, 2022.
Article in English | EMBASE | ID: covidwho-1820630

ABSTRACT

Excipients are critically important in converting active pharmaceutical ingredients (API) into drug products that have optimal stability, bioavailability, manufacturability, duration of action, and therapeutic benefits. They will play even greater roles in the future to enable drug targeting, delivery of biotech products and vaccines, gene therapy, continuous manufacturing, 3D printing, and so forth. This commentary describes the author’s experience in teaching a graduate course on excipients at St. John’s University to train students on optimal selection and appropriate use of excipients in formulating dosage forms and development of drug delivery systems. The course is offered in 15 two-hour sessions over a semester, and the course materials are divided into 13 modules on chemistry of different classes of polymeric and non-polymeric excipients and their application in dosage form development, including the use as solubilizing agents, lyophilizing agents, cryoprotectants, buffers, biodegradable materials, and carriers for amorphous solid dispersions and 3D printing. The development of coprocessed excipients, the need for new excipients, and the regulatory aspects of excipients are also covered. The course includes presentations by guest speakers from the industry, and the students also watch virtual presentations from experts that are publicly available from the internet. It is a popular course at St. John’s University taken by all graduate students in the pharmaceutics program. It is recommended that such courses are introduced in other pharmacy schools and academic institutions. The course may be adapted to meet specific needs of different academic programs. Professional associations, such as AAPS and CRS, industry groups like IPEC, and the pharmaceutical industry may be able to help in introducing such courses by providing lecture materials and guest lecturers.

19.
Embase; 2022.
Preprint in English | EMBASE | ID: ppcovidwho-333245

ABSTRACT

SARS-CoV-2 is a highly transmissible and pathogenic coronavirus that first emerged in late 2019 and has since triggered a pandemic of acute respiratory disease named COVID-19 which poses a significant threat to all public health institutions in the absence of specific antiviral treatment. Since the outbreak began in March 2020, India has reported 4.77 lakh Coronavirus deaths, according to the World Health Organization (WHO). The innate RNA interference (RNAi) pathway, on the other hand, allows for the development of nucleic acid-based antiviral drugs in which complementary small interfering RNAs (siRNAs) mediate the post-transcriptional gene silencing (PTGS) of target mRNA. Therefore, in this current study, the potential of RNAi was harnessed to construct siRNA molecules that target the consensus regions of specific structural proteins associated genes of SARS-CoV-2, such as the envelope protein gene (E), membrane protein gene (M), nucleocapsid phosphoprotein gene (N), and surface glycoprotein gene (S) which are important for the viral pathogenesis. Conserved sequences of 811 SARS-CoV-2 strains from around India were collected to design 21 nucleotides long siRNA duplex based on various computational algorithms and parameters targeting E, M, N and S genes. The proposed siRNA molecules possessed sufficient nucleotide-based and other features for effective gene silencing and BLAST results revealed that siRNAs' targets have no significant matches across the whole human genome and hence, siRNAs were found to have no off-target effects on the genome, ruling out the possibility of off-target silencing. Finally, out of 157 computationally identified siRNAs, only 4 effective siRNA molecules were selected for each target gene which is proposed to exert the best action based on GC content, free energy of folding, free energy of binding, melting temperature, heat capacity and molecular docking analysis with Human AGO2 protein. Our engineered siRNA candidates could be used as a genome-level therapeutic treatment against various sequenced SARS-CoV-2 strains in India. However, future applications will necessitate additional validations in vitro and in vivo animal models.

20.
Molecules ; 27(6)2022 Mar 17.
Article in English | MEDLINE | ID: covidwho-1760783

ABSTRACT

Shigella species account for the second-leading cause of deaths due to diarrheal diseases among children of less than 5 years of age. The emergence of multi-drug-resistant Shigella isolates and the lack of availability of Shigella vaccines have led to the pertinence in the efforts made for the development of new therapeutic strategies against shigellosis. Consequently, designing small-interfering RNA (siRNA) candidates against such infectious agents represents a novel approach to propose new therapeutic candidates to curb the rampant rise of anti-microbial resistance in such pathogens. In this study, we analyzed 264 conserved sequences from 15 different conserved virulence genes of Shigella sp., through extensive rational validation using a plethora of first-generation and second-generation computational algorithms for siRNA designing. Fifty-eight siRNA candidates were obtained by using the first-generation algorithms, out of which only 38 siRNA candidates complied with the second-generation rules of siRNA designing. Further computational validation showed that 16 siRNA candidates were found to have a substantial functional efficiency, out of which 11 siRNA candidates were found to be non-immunogenic. Finally, three siRNA candidates exhibited a sterically feasible three-dimensional structure as exhibited by parameters of nucleic acid geometry such as: the probability of wrong sugar puckers, bad backbone confirmations, bad bonds, and bad angles being within the accepted threshold for stable tertiary structure. Although the findings of our study require further wet-lab validation and optimization for therapeutic use in the treatment of shigellosis, the computationally validated siRNA candidates are expected to suppress the expression of the virulence genes, namely: IpgD (siRNA 9) and OspB (siRNA 15 and siRNA 17) and thus act as a prospective tool in the RNA interference (RNAi) pathway. However, the findings of our study require further wet-lab validation and optimization for regular therapeutic use for treatment of shigellosis.


Subject(s)
Dysentery, Bacillary , Shigella , Child , Diarrhea/drug therapy , Dysentery, Bacillary/drug therapy , Dysentery, Bacillary/genetics , Humans , RNA Interference , RNA, Small Interfering/metabolism , Shigella/genetics
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